The goals this Phase I SBIR grant are focused on the development of PG 301029 and chemically related compounds as therapeutics for treatment of hepatitis C virus infection. PG 301029 is a small molecule inhibitor of hepatitis C virus (HCV) replication that has been shown to be less toxic and have a larger therapeutic index than the approved therapeutic ribavirin in in vitro assays using the HCV surrogate bovine viral diarrhea virus (BVDV). The relative decrease in toxicity of PG 301029 compared to that of ribavirn was also observed when studies were extended to include a panel of primary and established cell lines, suggesting the potential of PG 301029 as a safer and more efficacious drug than ribavirin for the treatment of HCV. Additionally, preliminary in vivo toxicology studies indicate that the compound is well tolerated and has a pharmacokinetic profile appropriate for drug development. We will perform a structure activity relationship (SAR) based examination of an additional fifteen compounds previously identified as inhibitors of BVDV replication in a screen of 155 near neighbor related compounds for their ability to inhibit replication of an HCV replicon. These studies will provide insight into the functional structural features of PG 301029 which contribute to the antiviral activity and will be the basis for chemi- informatic studies to develop second generation PG 301029 based inhibitors as a component of the Phase II SBIR proposal. PUBLIC HEALTH RELEVANCE: The goal of this Phase I SBIR is to identify and characterize novel inhibitors of hepatitis C virus replication. The lead compound for development, PG 301029, is a small molecule that acts through a novel mechanism of action to reduce viral RNA accumulation in BVDV-infected and HCV replicon and has a therapeutic index greater >500 against BVDV. Structure activity relationship (SAR) analysis with 15 chemically related molecules that have shown antiviral activity against BVDV will be performed using a HCV replicon cell line to define the structural features of PG 301029 which contribute to their functional antiviral activity . The collective data will be the basis for chemi-informatic studies to develop second generation inhibitors as a component of the Phase II SBIR proposal. We will also use hypothesis directed research to define the mechanism of action of the antiviral activity by examining alterations in macromolecular interactions and gene expression in PG 301029 treated cells.